Novel Tetragenococcus halophilus with high yield of umami peptide and application of Tetragenococcus halophilus
A halophilic tetrad, umami peptide technology, applied in the direction of bacteria, microorganism-based methods, food ingredients as taste improvers, etc., can solve the problem of high cost, low yield of production strains, and inability to meet the needs of large-scale industrial production, etc. problems, to achieve the effect of short production cycle, high output and improved safety
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Embodiment 1
[0030] The isolation and screening of embodiment 1 bacterial strain
[0031] 1. Screening samples
[0032] Soybean paste collected from Shangpingzhou Village, Xiaotun Town, Liaoyang City, which has been naturally fermented for half a year.
[0033] 2. Preliminary screening of tetradococci
[0034] Take 1g of soybean paste sample, add 9mL of 0.9% sterile normal saline, mix well and soak for 15min; then take 1mL of sample solution and inoculate it in the enrichment medium, culture it at 37℃ for 1-2d, and spread it on a flat plate Cultured on solid separation medium for 5-6d.
[0035] Pick a single colony that is small, has obvious calcium-dissolving circle, is milky white and opaque, and has a shape similar to that of a lactic acid bacteria colony, and performs Gram staining. Microscopic observation results showed that there were 12 bacterial strains in the form of pairs or quadruple globules, which were named LY9-1, LY9-2, ..., LY9-12 respectively.
[0036] Purify and cultu...
Embodiment 2
[0053] Identification of embodiment 2LY9-1 bacterial strain
[0054] 1. Identification of colony morphology
[0055] The bacterium colony of LY9-1 bacterial strain described in the present invention is less, protrudes, and is milky white, opaque, and the surface is smooth and glossy, and the edge is flat; No motility, does not produce spores; Gram staining is positive, and the cell is spherical, quadruple or in pairs. Colony morphology and cell morphology figure 1 and figure 2 shown.
[0056] 2. Molecular biological identification
[0057] (1) Bacterial DNA extraction:
[0058] Pipette 100 μL of the LY9-1 bacterial suspension into the sterilized MRS liquid medium, and culture it in a shaking incubator at 37°C for 24 hours. The genome of the strain was extracted according to the operating steps of the bacterial genome kit (Solarbio D1600).
[0059] (2) PCR amplification:
[0060] PCR upstream primer 27F and downstream primer 1492R were designed and synthesized by Shang...
Embodiment 3
[0073] Example 3 Determination of Enzyme Production Ability of Tetradynococcus halophilus SNTH-1
[0074] 1. Determination of protease activity:
[0075] After the halophilic Tetradendococcus SNTH-1 strain was activated, it was inoculated in the MRS liquid medium and cultured in a constant temperature incubator at 37°C for 24 hours.
[0076] Prepare protease screening medium (10g casein, 3g yeast powder, 5g Na 2 HPO 4 12H 2O, 50g NaCl, 0.05g bromothymol blue, 20g agar, 1L distilled water, adjust the pH value to 7), and adopt the Oxford cup method to measure the protease-producing ability of the bacterial strain. Put the sterilized Oxford cup on the protease screening medium, absorb 10 μ of the halophilic Tetradendococcus SNTH-1 bacteria solution into the Oxford cup, and cultivate it in a constant temperature incubator at 37°C for 24 hours, and observe whether there is a transparent circle.
[0077] The casein hydrolysis circle diameter (D) and the colony diameter (d) were ...
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